The high mutation rates inherent to HIV have created sufficient sequence diversity to pose a formidable challenge in designing a universal HIV vaccine. Over the years since the start of the HIV epidemic, the virus has successfully utilized many viral and host pathways to avoid immune recognition. One of the best-characterized and most frequent viral adaptations is escape from cytotoxic T lymphocyte (CTL). This evasion method generates adapted epitopes (AE) many of which are not expected to be immunogenic since these are by-products of the CTL escape. These HLA-I restricted adaptations or sequence mutations in CTL epitopes appear to accumulate in a viral population and amplify over time if there is no pressure to revert; however, many revert rapidly due to viral fitness constraints when transmitted to new hosts who cannot target the specific epitope. The make up of a virus in a population is therefore in constant flux depending on the HLA-I alleles in that population driving escape and viral fitness constraints that force reversions back to its non- adapted state. Our preliminary data shows that a large proportion of CTL epitopes in a transmitted founder virus (TFV or virus that established infection in a new host) are already mutated with respect to that person's HLA-I (i.e. AE) making it unlikely that an effective and broad CD8 T-cell response will be elicited. In addition, we also have nascent data demonstrating a direct correlation between number of adapted epitopes (AE) transmitted during acute infection and set point viral load. Taken together, these observations highlight a truly concerning scenario pertinent directly to HIV vaccine design since all current HIV vaccines encode a large number of AE. To address this problem, we will first identify HIV specific CTL epitopes that would strongly merit inclusion n future candidate HIV immunogens. We hypothesize that a majority of these will be the non-adapted epitopes (NAE) which a) will effectively kill HIV infected cells; b) undergo CTL escape (to become AE); c) the escape inflicting a significant fitness cost to the virus; and d) the NAE reverts to AE when the immune pressure is lifted. These features and additional functional attributes proposed for evaluation in aim 1 will help delineate a signature for optimal CTL epitopes. Aim 2 will determine whether vaccines, used in prior efficacy studies, were enriched in AE thereby negating any effect on clinical markers of disease progression in vaccinees who became infected. We will also evaluate the immunogenicity of NAE and their AE counterparts in a candidate mosaic vaccine clinical trial. Finally, in aim 3 we gain mechanistic insights into how optimal CTL responses are generated by determining whether the structure of HLA-I bound peptides predicts the quality of the immune response. In summary, this proposal will define important non-adapted and adapted HIV epitopes in context of acute infection and vaccination. The information obtained will be immensely useful both for the design and generation of an evaluable benchmark for gauging the efficacy of future candidate HIV-1 vaccines.